guanosine-triphosphate and Hyperparathyroidism

The present studies were designed to examine the consequences of chronic mild elevations of endogenous parathyroid hormone (PTH) in vivo on the PTH receptor-adenylate cyclase system of canine kidney cortex. Hyperparathyroidism was induced in normal dogs by feeding a diet low in calcium, high in phosphorus to the animals for a period of 6-9 wk. This maneuver resulted in a two to threefold increase in the plasma levels of carboxy-terminal immunoreactive PTH. This degree of hyperparathyroidism is similar to that seen in patients with hyperparathyroidism and normal renal function. After 6-9 wk on the diet the animals were killed and basolateral renal cortical membranes prepared for the study of the PTH receptor-adenylate cyclase system in vitro. The dietary hyperparathyroidism resulted in desensitization of the PTH-responsive adenylate cyclase (Vmax 3,648 +/- 654 pmol cyclic (c)AMP/mg protein per 30 min in hyperparathyroid animals vs. 5,303 +/- 348 in normal controls). The Kact (concentration of PTH required for half-maximal enzyme activation) was unchanged. However, PTH receptor binding (125I-norleucyl8-norleucyl18-tyrosinyl34, 125I[Nle8, Nle18, Tyr34] bPTH (1-34) NH2 as radioligand) was not different in the two groups of animals. Thus, dietary hyperparathyroidism resulted in an uncoupling of the PTH receptor-adenylate cyclase system. This defect was not corrected by guanyl nucleotides in vitro, and the effects of guanyl nucleotides on PTH binding and enzyme activation appeared normal. NaF-stimulated enzyme activity was reduced in the hyperparathyroid animals (8,285 +/- 607 pmol cAMP/mg protein per 30 min vs. 10,851 +/- 247 in controls). These data indicate that desensitization of the PTH-responsive adenylate cyclase system of canine kidney as a result of mild chronic elevations of endogenous PTH is due to a postreceptor defect, demonstrable by NaF activation, not corrected by guanyl nucleotides, leading to abnormal PTH-receptor adenylate cyclase coupling.

Topics: Adenylyl Cyclases; Animals; Calcium; Diet; Dogs; Guanosine Triphosphate; Hyperparathyroidism; Kidney Cortex; Manganese; Phosphorus; Receptors, Cell Surface; Receptors, Parathyroid Hormone; Sodium Fluoride

A method of measuring the biological activity of parathyroid hormone (PTH) in human serum that depends on the activation of its natural target enzyme, human renal cortical adenylate cyclase, is described. Optimal sensitivity ranging in different assays from 14 to 20 pg 1-34 hPTH/ml was achieved in the presence of the GTP-analogue GppNHp (10 mumol/L), 5 mmol/L MgCl2 and 1.25 mmol/L EGTA. Basal and stimulated cAMP production was reproducible within assays (c.v. below 7%, S.E.M., n = 3) and between assays (c.v. 5 to 14%, S.E.M., n = 4). The recovery of 1-34 hPTH added to individual test sera averaged 94%. The specificity of the method was established as follows: 1.) Other tested hormones, at 100 ng/ml, were ineffective; 2.) In the majority of peripheral sera from patients with hyperfunctioning parathyroid glands elevated bio-activity was detected; 3.) The circulating bio-activity fell rapidly after removal of parathyroid adenomata; 4.) Treatment with antisera for hPTH reduced the bio-activity; 5.) A PTH-antagonist inhibited the bio-activity.

Topics: Adenoma; Adenylyl Cyclases; Biological Assay; Guanosine Triphosphate; Humans; Hyperparathyroidism; Kidney Cortex; Parathyroid Hormone; Parathyroid Neoplasms

Current evidence suggests that parathyroid gland adenylate cyclase is involved in the control of parathyroid hormone (PTH) secretion. Thus, the altered control of PTH release in hyperparathyroidism may relate to altered adenylate cyclase activation. Therefore, we examined adenylate cyclase kinetics in membrane preparations from hyperfunctioning human parathyroid glands and normal human and bovine parathyroid tissues. There were no differences in the affinity for ATP between enzymes of normal and pathological tissue. However, the enzyme in 10 hyperfunctioning glands showed increased affinity for Mg++. The activation constant for Mg++ (KaMg) of adenylate cyclase in normal human glands was 10.6 +/- 2 mM, a value not different from that of normal bovine parathyroid tissue (9.5 +/- 1 mM). In contrast, the adenylate cyclase in membrane preparations from three of four hyperplastic and six of seven adenomatous human glands showed a markedly reduced KaMg, ranging from 0.85-1.64 mM and from 1.58-6.46 mM, respectively. In one adenoma and one hyperplastic gland, the Ka of the enzyme for Mg++ was close to normal. The addition of guanylylimidodiphosphate or GTP to the incubation mixture increased, in a dose-dependent manner, the apparent KaMg of the enzyme in the abnormal tissue toward normal, suggesting a defective nucleotide regulatory site in the adenylate cyclase of hyperparathyroid glands. In addition, the hyperparathyroid gland enzyme was less susceptible to inhibition by calcium, requiring 0.7-1 mM Ca++ for 50% inhibition, whereas comparable inhibition of the normal adenylate cyclase was seen at 0.22-0.28 mM Ca++. We conclude that the abnormal control of PTH secretion in hyperparathyroidism may be related, at least in part, to alterations in the characteristics of parathyroid gland adenylate cyclase.

Topics: Adenosine Triphosphate; Adenylyl Cyclases; Animals; Calcium; Cattle; Guanosine Triphosphate; Guanylyl Imidodiphosphate; Humans; Hyperparathyroidism; Kinetics; Magnesium; Parathyroid Glands